Multiple myeloma (MM) is an incurable hematological cancer characterized by MM cells accumulation in the bone marrow (BM) that promotes tumor survival, drug resistance and consequent patient's relapse. The oncogenic Notch pathway consists of 4 receptors (Notch1-4) activated upon binding of two families of ligands, Jag (Jag1 and 2) and Dll (Dll1, 3 and 4) and plays a crucial role in MM. The aberrant expression of Notch receptors and ligands in tumor cell results in homotypical and heterotypical interactions with the BM that further favours tumor progression, osteoclastogenesis and drug resistance. In particular, aberrant Notch2 activation and Jag2 overexpression in MM cells play an important role in MM progression by stimulating osteoclast differentiation, release of pro-tumor cytokines by BM niche cells and MM cell self-renewal. Therefore uncoupling the interaction between Notch2 receptor and Jag2 ligand is critical to affect not only MM cells growth, but also their interaction with the BM niche. AIM: These evidences prompt us to develop a therapeutic tool to selectively inhibit Notch2 signaling triggered by Jag2 using an unprecedented approach based on druglike small molecules. MATERIAL AND METHODS. Currently, indirect approaches to inhibit Notch signaling are mainly based on inhibition of -Secretase that catalyzes Notch activation and thereby affect also other -Secretase substrates. Moreover, inhibition of all four Notch receptors is associated with gut toxicity that might be avoided by selectively blocking Notch signaling triggered by only one family of ligands, Jag or Dll ligands. Therefore, to selectively target Notch signaling members dysregulated in MM, we applied an inhibitory strategy based on small molecules that are metabolically stable, orally active and can be easily delivered. We have used in silico protein-protein docking and virtual high-throughput screening (HTS) of an Asinex chemoteque to select small molecules. The biological activity was validated through a Notch responsive gene reporter assay and two-cell co-culture assay that allows to measure Notch transcriptional activity induced either by Dll or Jag ligands. RESULTS: We have set-up a strategy to exclusively uncouple Notch2::Jag2, leaving unaltered the interaction with Dll. To this, we refined an integrated in silico pipeline already set up in the laboratory that brought us to identification of two small molecules with anti-proliferative efficacy and able to reduce anti-Notch transcriptional activity. A new approach took advantage of the differences in the surfaces of Notch2::Jag2 and Notch2::Dll4 complexes, modelled by protein::protein. This allowed us to select in silico 100 top-scoring compounds directed exclusively to Notch2::Jag2 surface by HTS of chemoteque of druglike small molecules. Preliminary, 2 of 100 compounds were validated in vitro. A Notch responsive reporter assay on HEK293T cells showed that compounds were able to significantly reduce Notch transcriptional activity. A Notch reporter assay of co-culture systems to measure Notch2 activation triggered either by Dll4 or Jag1/2 ligands detected that one of two compounds more specifically inhibited Notch2::Jag1/2 but not Notch2::Dll4 interactions. Our integrated pipeline represents a successful strategy to identify compounds that directly and selectively antagonize Notch activation and lays a basis for the development of entirely novel drugs to inhibit Notch signaling in cancer.
New Notch-targeted therapeutic strategy to uncouple a pathological interaction of myeloma cells and bone marrow microenvironment using novel small molecules / N. Platonova, C. Parravicini, L. Palazzolo, S. Saporiti, M. Colombo, V. Vallelonga, D. Colella R, B. F, A. Neri, I. Eberini, R. Chiaramonte. ((Intervento presentato al convegno Congresso DISS tenutosi a Milano nel 2017.
|Titolo:||New Notch-targeted therapeutic strategy to uncouple a pathological interaction of myeloma cells and bone marrow microenvironment using novel small molecules|
PLATONOVA, NATALIA (Primo)
PARRAVICINI, CHIARA (Secondo)
|Data di pubblicazione:||13-nov-2017|
|Settore Scientifico Disciplinare:||Settore MED/04 - Patologia Generale|
Settore BIO/13 - Biologia Applicata
|Citazione:||New Notch-targeted therapeutic strategy to uncouple a pathological interaction of myeloma cells and bone marrow microenvironment using novel small molecules / N. Platonova, C. Parravicini, L. Palazzolo, S. Saporiti, M. Colombo, V. Vallelonga, D. Colella R, B. F, A. Neri, I. Eberini, R. Chiaramonte. ((Intervento presentato al convegno Congresso DISS tenutosi a Milano nel 2017.|
|Appare nelle tipologie:||14 - Intervento a convegno non pubblicato|